The present invention relates to mass spectrometry apparatus and methods for obtaining multidimensional data which describe relationships between parent ions and daughter ions produced by fragmentation, such as has been previously obtained by tandem mass spectrometers. The invention further relates to multidimensional analysis techniques for improving mass resolution in single-stage mass spectrometers.
In simple mass spectrometers, sample ions are formed such as by electron ionization (EI), passed through a mass analyzer such as a magnetic sector, and detected. The detected ions can be molecular ions, fragment ions of the molecular ion, or fragment ions of other fragments.
Selected ion fragmentation mass spectrometers have recently been developed, characterized by having two sequential stages of mass analysis and an intermediate fragmentation region. Hence, these are generally termed "tandem" or "MS/MS" instruments. In such tandem mass spectrometers, sample ions are produced in an ion source, and the first stage of mass analysis selects parent ions of particular mass. Then, some of the selected parent ions fragment or dissociate, such as by metastable decomposition, collision induced dissociation (CID), or collisionally activated dissociation (CAD), producing daughter ions. Finally, the second stage of mass analysis selects the parent ion and its daughter ions according to mass.
These instruments provide the ability to identify parent ions and the daughter ions which result from fragmentation. The complete MS/MS spectrum is a multidimensional spectrum showing all the daughter ions of each of the parent ions. Subsets of the complete MS/MS data such as a spectrum showing all the daughters of a specific parent ion (daughter spectra) are proving to be invaluable for many applications in complex mixture analysis and structure elucidation. Also proving invaluable are spectra which show all the parents of a particular daughter mass (parent spectra). Useful, too, are spectra which show all parent ions which lose a particular mass during fragmentation, known as neutral loss spectra. MS/MS is an extremely useful technique due to the large amount of characterizing data which may be obtained from a single sample by collecting particular parent ion, daughter ion, or neutral loss spectra, or by collecting the complete MS/MS spectrum.
Heretofore, there have been two general types of tandem mass spectrometers for obtaining MS/MS data. The first of these general types of instruments is a double-focusing or double-sector instrument, a common one being of the Mass-Analyzed Ion Kinetic Energy Spectrometry (MIKES) type. In a MIKES-type instrument, a magnetic momentum selector (magnetic sector) and an electrostatic kinetic energy selector (electric sector) are coupled in tandem, with a fragmentation region between the two sectors. The magnetic sector selects parent ions of particular momentum (related to mass) for fragmentation. The electrostatic sector then produces an ion kinetic energy separation which is interpreted to provide the fragmentation mass spectrum. Such an instrument may be realized with commercially available reversed-geometry double-focusing mass spectrometers.
Both magnetic and electric (electrostatic) sector instruments operate by ion path bending under the influence of the magnetic or electric field, as the case may be. The path radius or, stated alternatively, the deflection angle, is a function of both mass and field strength, as well as of ion velocity. In scanning type magnetic or electrostatic mass spectrometers, ion groups of differing mass are successively swept through a single slit by varying magnetic or electrostatic field strength as a function of time. Thus, in any moment, only a single beam exits the slit, and registers as ion current in an ion detector, typically an electron multiplier.
The second general type of tandem mass spectrometer is known as a Triple Quadrupole Mass Spectrometer (TQMS), so termed because two quadrupole mass analyzers are employed to respectively select parent ions and daughter ions and a third, intermediate, quadrupole operated in the RF-only mode comprises the collision chamber. A triple quadrupole mass spectrometer is disclosed in Enke et al U.S. Pat. No. 4,234,791.
A quadrupole is an electrodynamic focusing device including DC and RF electric fields and which operates as a mass filter. Only ions of selected mass pass through the device, the selected mass being a function of the DC and RF electric fields.
Compared to MIKES, TQMS instruments have a number of significant advantages, including high sensitivity permitting use in trace analysis, rapid scanning speeds permitting relatively high rates of data acquisition when coupled to computer-based control systems, and improved mass resolution for daughter ions over that possible with magnetic and electric sectors as employed in MIKES. The MIKES-type instruments have the advantage of a greater mass range compared to TQMS instruments.
A fundamental disadvantage of tandem mass spectrometers, both of the MIKES-type and the TQMS-type, results from the fact that in operation they are sequential in nature. Only one combination of parent and daughter ion masses is within the ion flight path within the instrument at any one time. To obtain a complete MS/MS fragmentation spectrum showing all possible relationships between parent ions and daughter ions requires that the second mass analyzer (which selects daughter ions) be completely scanned for each potential parent isolated by the first mass analyzer (which selects parent ions). While the quadrupole-based tandem mass spectrometer (TQMS) scans faster than MIKES, the sequential nature of its operation nevertheless limits the rates at which spectral data can be acquired.
Tandem mass spectrometers have been combined with gas chromatographs and liquid chromatographs, resulting in instruments which are respectively termed "GC-MS/MS" and "LC-MS/MS". However, MS/MS heretofore has been too slow to obtain a full multidimensional MS/MS spectrum during the relatively brief period (e.g. one to ten seconds) of a chromatographic peak. Rather, GC-MS/MS is currently implemented for the purpose of selected reaction monitoring (for a particular parent-daughter combination) in which both mass filters are either set at fixed mass numbers, or scan a limited mass range. Ions selected by the first mass filter undergo collision, and selected products of the fragmentation reaction are monitored. Clearly only limited information about the sample is obtained with this technique.
While the foregoing summarizes the two most important instrument types for MS/MS, in the particular context of the present invention there are several other types of mass spectrometers which deserve mention.
As an alternative to magnetic or electric field scanning, spatial array detectors have been proposed which are similar in concept to the traditional use of a strip of photographic film as the output detector of a mass spectrograph, but have the advantage of providing output data in real time as electrical signals. For example, microchannel electron multiplier arrays (MCA) have been proposed, as well as electro-optical ion detectors (EOID) such as is described in Giffin U.S. Pat. No. 3,955,084.
Also deserving mention in the context of the present invention is another form of mass spectrometry known as time-of-flight (TOF) mass spectrometry. TOF mass spectrometry does not rely on path bending as in magnetic or electrostatic spectrometers, nor on electrodynamic filtering of the type which occurs in a quadrupole-type mass filter.
Briefly, in a time-of-flight mass spectrometer ions are produced and then accelerated, either in a constant-energy or a constant-momentum mode.
In either case, lighter (lower mass) ions are accelerated to higher velocities than the heavier ions. The ions then enter a drift region or flight tube which establishes an ion path length, and which is followed by an ion detector. In the drift region, the ions separate along the ion path as a function of their velocity and thus arrive at the detector at different times depending upon their velocities, and therefore, depending upon their mass.
To permit measurement of flight time, ions in a time-of-flight mass spectrometer are bunched, typically by means of a pulsed source, and all ions of a given bunch enter the drift region at substantially the same position and time. By correlating ion pulsing or bunching with arrival time of various ions at the detector, the time-of-flight of each individual ion or group of identical-mass ions can be determined. Ion velocity follows from the simple relationship: (Velocity)=(Path Length)/(Time-Of-Flight). From velocity, ion mass can be calculated, taking into account the characteristics of the ion accelerator.
Time-of-flight mass spectrometry is known to have a number of advantages, including an extremely fast scanning or cycle rate (typically ten thousand mass spectra per second), and potentially unlimited mass range.
Commonly-available commercial time-of-flight instruments measure detected ion current intensities by sampling techniques. Ion current is sampled during only one arrival time for each source pulse. A sampling window or time slice (aperture time) is established, defined by a time delay from the source pulse to the window. The delay time is slowly scanned while the source is repetitively pulsed. A complete mass spectrum of the sample under study is recorded by collecting the ion intensities for each successive arrived time. This technique is known as Time-Slice Detection (TSD).
Also, in order to obtain a statistically valid number of samples, integrating forms of time-slice detectors have been employed, known as "boxcar integrators". The boxcar integrator is triggered for each ion pulse, and integrates ion current during the same aperture time at a constant arrival time for a number of pulses. The aperture time can either be constant or be slowly scanned.
Time-Slice Detection has the disadvantage of losing most of the information available in the ion beam since aperture time is a small fraction of the total time over which ions are arriving at the detector. This creates a potential problem where sample quantities are limited. Accordingly, various devices for Time Array Detection (TAD) have been proposed, known variously as "transient recorders" or "digital transient recorders". Such recorders, rather than responding to a single time slice relative to the pulsed source, collect the entire output from a single source pulse in a time-of-flight mass spectrometer to produce individual data channels (or "time bins") for each of a multiplicity of sample points taken serially in time.
Heretofore available digital transient recorders, although offering an improvement over time-slice detection, are not capable of measuring ten thousand transients per second consistent with the ten thousand per second pulse rate typical in TOF mass spectrometry, and thus lose data as a result of spectra not collected. In particular, their data readout time is in the order of milliseconds, and is inconsistent with the 10 kHz or greater pulse rate of time-of-flight analysis.
Just as a boxcar integrator is an integrating form of time-slice detector, integrating forms of digital transient recorders have been employed, although operating relatively slowly. One name for such devices is Computer of Average Transients, or "CAT". Another form of integrating transient recorder, reported by Linclon, comprises a Biomation Model 8100 digital transient recorder compled to a Nicolet Model 1170 signal averager. See K. A. Linclon, "Data Acquisition Techniques for Exploiting the Uniqueness of the Time-of-Flight Mass Spectrometer: Application to Sampling Pulsed Gas Systems" Dyn. Mass, Spectrom., 6, 111-119 (1981); also published as NASA Report Tm-81224.
An advanced form of integrating transient recorder operating at the required data rate is disclosed in commonly-assigned U.S. patent application Ser. No. 385,115, filed June 1, 1982, concurrently herewith, by Christie George ENKE, Bruce Hewitt NEWCOME and John Francis HOLLAND and entitled "HIGH REPETITION RATE TRANSIENT RECORDER WITH AUTOMAIC INTEGRATION."
Resolution in TOF mass spectrometry would be limited only by detector response speed if all ions started from an initial plane with zero initial energy. This is not the case in reality. There have been a number of efforts to increase resolution in TOF instruments by special focusing techniques. For example, the object of "energy focusing" is to render the produced mass spectrum independent of initial ion kinetic energy. The object of "momentum foucusing" is to render the produced mass spectrum independent of initial ion momentum. More generally, the object of "velocity focusing" is to render the produced mass spectrum independent of initial ion velocity. Similiarly, the object of "space focusing" is to compensate for the initial space distribution. For example, see Wiley and McLaren, Rev. Sci. Instrum., 26, 1150-1157 (1955). Another effort has been the use of V-shaped and linear reflection devices, which also operate on a space focusing principle. For example, see B. A. Mamyrin, V. I. Karataeu, P. V. Shmikk, V. A. Zagulin, Sov. Phys JETP, 37, 45-48 (1973); and B. A. Mamyrin, D. V. Shmikk, Sov. Phys. JETP, 49, 762-764 (1979).
In the particular context of the present invention, it is also pertinent to note that electric sectors for "energy focusing" and magnetic sectors for "momentum focusing" have been proposed for enhancing the revolution of TOF mass analysis. See, for example, Moorman et al U.S. Pat. No. 3,576,992 and Poschenrieder U.S. Pat. No. 3,863,068.